Project Summary Heart failure in elderly and/or hypertensive patients is a major public health epidemic that predisposes to sudden cardiac death. Fibrosis is a hallmark feature of these patients that contributes significantly to arrhythmogenesis. Existing therapies for combatting arrhythmias in heart failure patients are inadequate in large part because they fail to reverse preexisting fibrosis. The matricellular protein CCN5 mediates disease regression by eliciting an adaptive cardiac response against pressure overload induced stress. While endogenous CCN5 exerts a potent anti-fibrosis efficacy, we found that chronic AAV9-mediated overexpression (OE) of CCN5 in models of advanced heart failure with increased afterload (HF/iAL) that mimic the effects of aging and/or hypertension failed to restore conduction or prevent arrhythmias. Furthermore, our preliminary data demonstrate that AAV9-mediated CCN5 OE causes adverse electrical remodeling that precedes structural remodeling in a chronic model of pressure overload. Finally, we found that AAV9-mediated CCN5 OE in structurally normal (fibrosis-free) hearts caused significant conduction slowing with no change in membrane excitability suggesting a hitherto unrecognized inhibitory effect of CCN5 on inter-cellular coupling that may interfere with its beneficial efficacy in the setting of advanced heart failure. Our central hypothesis is that the potential anti-arrhythmic efficacy of suppressing fibrosis through CCN5 is hampered by detrimental (fibrosis-independent) electrophysiological effects. Our current studies are tailored to define the fibrosis- dependent and independent properties of CCN5 and to harness this knowledge therapeutically. We aim to retain the beneficial and avoid the detrimental (fibrosis-independent) effects of CCN5 through rapid, local, and transient delivery using a chemically modified RNA gene delivery approach as opposed to chronic global transduction. Overall, completion of this project will lead to improved understanding of the role of fibrosis in arrhythmogenesis and guide the development of mechanism-based approaches for combatting arrhythmias in patients with advanced heart failure who currently have very limited therapeutic options. These studies may further establish a novel mechanism by which cell-to-cell coupling is regulated in health and disease. Studies in this high-risk R21 proposal address critical barriers that currently obstruct a potentially power anti-fibrosis therapy from advancing forward.